Analysis of MIF,FCGR2A and FCGR3A gene polymorphisms with susceptibility to pulmonary tuberculosis in Moroccan population

In order to investigate the influence of functional polymorphisms of macrophage migration inhibitory factor(MIF),Fcg receptors CD16A(FCGR3A) and CD32A(FCGR2A) genes on susceptibility to pulmonary tuberculosis(PTB) in the Moroccan population,we analyzed 123 patients with PTB and 154 healthy controls.The genotyping for MIF-173(G/C)(rs755622),FCGR2A-131H/R(rs1801274) and FCGR3A-158V/F(rs396991) was carried out using TaqMan SNP Genotyping Assay method.We found a statistically significant increase of the MIF-173...     

Analysis of MIF,FCGR2A and FCGR3A gene polymorphisms with susceptibility to pulmonary tuberculosis in Moroccan population

In order to investigate the influence of functional polymorphisms of macrophage migration inhibitory factor(MIF),Fcg receptors CD16A(FCGR3A) and CD32A (FCGR2A) genes on susceptibility to pulmonary tuberculosis(PTB)in the Moroccan population,we analyzed 123 patients with PTB and 154 healthy controls.The genotyping for MIF-173(G/C)(rs755622),FCGR2A-131H/R(rsl801274)and FCGR3A-158V/F(rs396991) was carried out using TaqMan SNP Genotyping Assay method.We found a statistically significant increase of the MIF-173CC homozygote genotype and MIF-173*C allele frequencies in PTB patients compared with healthy controls (17.07%versus 5.84%,P=0.003;and 35.37%versus 26.30%,P=0.02;respectively).In contrast,no association was observed between CGR2A-131H/R and FCGR3A-158V/F polymorphisms and tuberculosis disease.Our finding suggests that MIF-173*C variant may play an important role in the development of active tuberculosis.     

Analysis of Fcgamma receptor haplotypes in rheumatoid arthritis: FCGR3A remains a major susceptibility gene at this locus, with an additional contribution from FCGR3B

The Fcgamma receptors play important roles in the initiation and regulation of many immunological and inflammatory processes, and genetic variants (FCGR) have been associated with numerous autoimmune and infectious diseases. The data in rheumatoid arthritis (RA) are conflicting and we previously demonstrated an association between FCGR3A and RA. In view of the close molecular proximity with FCGR2A, FCGR2B and FCGR3B, additional polymorphisms within these genes and FCGR haplotypes were examined to refine the extent of association with RA. Biallelic polymorphisms in FCGR2A, FCGR2B and FCGR3B were examined for association with RA in two well characterized UK Caucasian and North Indian/Pakistani cohorts, in which FCGR3A genotyping had previously been undertaken. Haplotype frequencies and linkage disequilibrium were estimated across the FCGR locus and a model-free analysis was performed to determine association with RA. This was followed by regression analysis, allowing for phase uncertainty, to identify the particular haplotype(s) that influences disease risk. Our results reveal that FCGR2A, FCGR2B and FCGR3B were not associated with RA. The haplotype with the strongest association with RA susceptibility was the FCGR3A-FCGR3B 158V-NA2 haplotype (odds ratio 3.18, 95% confidence interval 1.13-8.92 [P = 0.03] for homozygotes compared with all genotypes). The association was stronger in the presence of nodules (odds ratio 5.03, 95% confidence interval 1.44-17.56; P = 0.01). This haplotype was also more common in North Indian/Pakistani RA patients than in control individuals, but not significantly so. Logistic regression analyses suggested that FCGR3A remained the most significant gene at this locus. The increased association with an FCGR3A-FCGR3B haplotype suggests that other polymorphic variants within FCGR3A or FCGR3B, or in linkage disequilibrium with this haplotype, may additionally contribute to disease pathogenesis.     

Association of glutathione-S-transferase omega haplotypes with susceptibility to chronic obstructive pulmonary disease

Cigarette smoking is the main risk factor for developing the inflammatory lung disease chronic obstructive pulmonary disease (COPD). Differences in susceptibility among smokers have been attributed to a genetic predisposition. A recent publication on the Framingham Heart Study found a strong association of the Asn142Asp SNP in Glutatthione-S-transferase Omega (GSTO) 2 with forced expiratory volume in the first second (FEV₁) and forced vital capacity (FVC). FEV₁ is the main parameter reflecting the degree of airflow limitation in patients with COPD. Therefore the present study was undertaken to investigate whether the Asn142Asp polymorphism in GSTO2 occurs more frequently in patients with COPD than healthy subjects and to replicate the finding that it strongly correlates with FEV₁. Furthermore, the Ala140Asp substitution in GSTO1 was examined. Genotyping was carried out in 195 healthy controls and 355 patients with COPD. The results demonstrate that the Asn142Asp polymorphism in GSTO2 and the GSTO1140Asp/GSTO2142Asp haplotype were associated with increased risk of COPD. However, single-marker and haplotype-based analyses failed to reveal an association between lung function parameters and investigated non-synonymous coding SNPs in the GSTO genes. In conclusion, GSTO2 is a candidate gene for COPD, but is not associated with FEV₁.     

FCGR2C genotyping by pyrosequencing reveals linkage disequilibrium with FCGR3A V158F and FCGR2A H131R polymorphisms in a Caucasian population

The FCGR3A-V158F and FCGR2A-H131R polymorphisms are associated with clinical responses to therapeutic mAbs and with immune thrombocytopenic purpura (ITP). The FCGR2C-ORF/STOP polymorphism, controlling FcγRIIC expression on natural killer cells and therefore FcγRIIC-mediated antibody dependent cell-mediated cytotoxicity, is also associated with ITP. Using a new pyrosequencing assay to determine this polymorphism in a control population, we observed the expected allele frequencies (ORF:12.6%) and percentages of individuals with a single copy (10.0%) or 3 copies (12.1%) of FCGR2C, or with at least one FCGR2C-ORF allele (20.1%). No association of FCGR2C copy number variations with the FCGR3A-V158F or FCGR2A-H131R genotype was detected. More importantly, our results demonstrate a strong and a weaker linkage disequilibrium associating the FCGR2C-ORF allele with the FCGR3A-158V and the FCGR2A-131H allele, respectively.     

FCGR2C genotyping by pyrosequencing reveals linkage disequilibrium with FCGR3A V158F and FCGR2A H131R polymorphisms in a Caucasian population

The FCGR3A-V158F and FCGR2A-H131R polymorphisms are associated with clinical responses to therapeutic mAbs and with immune thrombocytopenic purpura (ITP). The FCGR2C-ORF/STOP polymorphism, controlling FcγRIIC expression on natural killer cells and therefore FcγRIIC-mediated antibody dependent cell-mediated cytotoxicity, is also associated with ITP. Using a new pyrosequencing assay to determine this polymorphism in a control population, we observed the expected allele frequencies (ORF:12.6%) and percentages of individuals with a single copy (10.0%) or 3 copies (12.1%) of FCGR2C, or with at least one FCGR2C-ORF allele (20.1%). No association of FCGR2C copy number variations with the FCGR3A-V158F or FCGR2A-H131R genotype was detected. More importantly, our results demonstrate a strong and a weaker linkage disequilibrium associating the FCGR2C-ORF allele with the FCGR3A-158V and the FCGR2A-131H allele, respectively.     

Associations between FCGR2A rs1801274, FCGR3A rs396991, FCGR3B NA1/NA2 polymorphisms and periodontitis: a meta-analysis

The aim of this study was to determine whether the Fcγ receptors (FCGRs) polymorphisms confer susceptibility to periodontitis in ethnically different populations. We did a literature search using PubMed and Embase, and conducted a meta-analysis on the associations between the FCGR2A H131R (rs1801274), FCGR3A F158V (rs396991), and FCGR3B NA1/NA2 polymorphisms and periodontitis using allele contrast, the recessive model, the dominant model, and the homozygote contrast. A total of 17 separate comparisons with 1,421 patients with periodontitis and 1,454 controls, involving six Caucasian, six East Asian, two African and one South Asian population were considered in the meta-analysis. Meta-analysis of the FCGR2A H131R polymorphism showed no association between periodontitis and the FCGR2A R allele (OR = 0.987, 95 % CI = 0.881–1.107, p = 0.827). Stratification by ethnicity revealed an association between the RR+RH genotype with periodontitis in Caucasian population (OR = 0.624, 95 % CI = 0.479–0.813, p = 4.7 × 10^?5), but not in East Asian, and African populations. Meta-analysis of the FCGR3A F158V polymorphism revealed an association between the FCGR3A V allele and periodontitis is in Caucasians (OR = 1.457, 95 % CI = 1.014–2.092, p = 0.042), but not in East Asians and Africans. In addition, analysis using the dominant model and homozygote contrast showed the same pattern for the FCGR3A V allele. Meta-analysis of the FCGR3B NA1/NA2 polymorphism using the recessive model revealed a significant association between the NA2/NA2 genotype and periodontitis in aggressive periodontitis (OR = 2.853, 95 % CI = 1.673–4.863, 1.1 × 10^?5). This meta-analysis demonstrates that the FCGR2A, and FCGR3A polymorphisms may confer susceptibility to periodontitis in Caucasians, and that the FCGR3B polymorphism may be associated with susceptibility to aggressive periodontitis.     

Association of HLA-DRB1 amino acid residues with giant cell arteritis: genetic association study,meta-analysis and geo-epidemiological investigation

IntroductionGiant cell arteritis (GCA) is an autoimmune disease commonest in Northern Europe and Scandinavia. Previous studies report various associations with HLA-DRB1*04 and HLA-DRB1*01; HLA-DRB1 alleles show a gradient in population prevalence within Europe. Our aims were (1) to determine which amino acid residues within HLA-DRB1 best explained HLA-DRB1 allele susceptibility and protective effects in GCA, seen in UK data combined in meta-analysis with previously published data, and (2) to determine whether the incidence of GCA in different countries is associated with the population prevalence of the HLA-DRB1 alleles that we identified in our meta-analysis.MethodsGCA patients from the UK GCA Consortium were genotyped by using single-strand oligonucleotide polymerization, allele-specific polymerase chain reaction, and direct sequencing. Meta-analysis was used to compare and combine our results with published data, and public databases were used to identify amino acid residues that may explain observed susceptibility/protective effects. Finally, we determined the relationship of HLA-DRB1*04 population carrier frequency and latitude to GCA incidence reported in different countries.ResultsIn our UK data (225 cases and 1378 controls), HLA-DRB1*04 carriage was associated with GCA susceptibility (odds ratio (OR) = 2.69, P = 1.5×10⁻¹¹), but HLA-DRB1*01 was protective (adjusted OR = 0.55, P = 0.0046). In meta-analysis combined with 14 published studies (an additional 691 cases and 4038 controls), protective effects were seen from HLA-DR2, which comprises HLA-DRB1*15 and HLA-DRB1*16 (OR = 0.65, P = 8.2×10⁻⁶) and possibly from HLA-DRB1*01 (OR = 0.73, P = 0.037). GCA incidence (n = 17 countries) was associated with population HLA-DRB1*04 allele frequency (P = 0.008; adjusted R² = 0.51 on univariable analysis, adjusted R² = 0.62 after also including latitude); latitude also made an independent contribution.ConclusionsWe confirm that HLA-DRB1*04 is a GCA susceptibility allele. The susceptibility data are best explained by amino acid risk residues V, H, and H at positions 11, 13, and 33, contrary to previous suggestions of amino acids in the second hypervariable region. Worldwide, GCA incidence was independently associated both with population frequency of HLA-DRB1*04 and with latitude itself. We conclude that variation in population HLA-DRB1*04 frequency may partly explain variations in GCA incidence and that HLA-DRB1*04 may warrant investigation as a potential prognostic or predictive biomarker.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0692-4) contains supplementary material, which is available to authorized users.     

Genomic Pathology of SLE-Associated Copy-Number Variation at the FCGR2C/FCGR3B/FCGR2B Locus

Reduced FCGR3B copy number is associated with increased risk of systemic lupus erythematosus (SLE). The five FCGR2/FCGR3 genes are arranged across two highly paralogous genomic segments on chromosome 1q23. Previous studies have suggested mechanisms for structural rearrangements at the FCGR2/FCGR3 locus and have proposed mechanisms whereby altered FCGR3B copy number predisposes to autoimmunity, but the high degree of sequence similarity between paralogous segments has prevented precise definition of the molecular events and their functional consequences. To pursue the genomic pathology associated with FCGR3B copy-number variation, we integrated sequencing data from fosmid and bacterial artificial chromosome clones and sequence-captured DNA from FCGR3B-deleted genomes to establish a detailed map of allelic and paralogous sequence variation across the FCGR2/FCGR3 locus. This analysis identified two highly paralogous 24.5 kb blocks within the FCGR2C/FCGR3B/FCGR2B locus that are devoid of nonpolymorphic paralogous sequence variations and that define the limits of the genomic regions in which nonallelic homologous recombination leads to FCGR2C/FCGR3B copy-number variation. Further, the data showed evidence of swapping of haplotype blocks between these highly paralogous blocks that most likely arose from sequential ancestral recombination events across the region. Functionally, we found by flow cytometry, immunoblotting and cDNA sequencing that individuals with FCGR3B-deleted alleles show ectopic presence of FcγRIIb on natural killer (NK) cells. We conclude that FCGR3B deletion juxtaposes the 5′-regulatory sequences of FCGR2C with the coding sequence of FCGR2B, creating a chimeric gene that results in an ectopic accumulation of FcγRIIb on NK cells and provides an explanation for SLE risk associated with reduced FCGR3B gene copy number.     

Neurotrophins are expressed in giant cell arteritis lesions and may contribute to vascular remodeling

Introduction

Giant cell arteritis (GCA) is characterized by intimal hyperplasia leading to ischaemic manifestations that involve large vessels. Neurotrophins (NTs) and their receptors (NTRs) are protein factors for growth, differentiation and survival of neurons. They are also involved in the migration of vascular smooth muscle cells (VSMCs). Our aim was to investigate whether NTs and NTRs are involved in vascular remodelling of GCA.

Methods

We included consecutive patients who underwent a temporal artery biopsy for suspected GCA. We developed an enzymatic digestion method to obtain VSMCs from smooth muscle cells in GCA patients and controls. Neurotrophin protein and gene expression and functional assays were studied from these VSMCs. Neurotrophin expression was also analysed by immunohistochemistry in GCA patients and controls.

Results

Whereas temporal arteries of both GCA patients (n = 22) and controls (n = 21) expressed nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), tropomyosin receptor kinase B (TrkB) and sortilin, immunostaining was more intense in GCA patients, especially in the media and intima, while neurotrophin-3 (NT-3) and P75 receptor (P75^NTR) were only detected in TA from GCA patients. Expression of TrkB, a BDNF receptor, was higher in GCA patients with ischaemic complications. Serum NGF was significantly higher in GCA patients (n = 28) vs. controls (n = 48), whereas no significant difference was found for BDNF and NT-3. NGF and BDNF enhanced GCA-derived temporal artery VSMC proliferation and BDNF facilitated migration of temporal artery VSMCs in patients with GCA compared to controls.

Conclusions

Our results suggest that NTs and NTRs are involved in vascular remodelling of GCA. In GCA-derived temporal artery VSMC, NGF promoted proliferation and BDNF enhanced migration by binding to TrkB and p75^NTR receptors. Further experiments are needed on a larger number of VSMC samples to confirm these results.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-014-0487-z) contains supplementary material, which is available to authorized users.     

Messages from the morgue to rheumatology: from Chagas disease to giant cell arteritis

Autopsy rates have fallen from over 50% to less than 10% in recent decades. The drastic decline creates uncertainty regarding causes of death and has negative effects for research, training, and patient safety, despite advances in imaging and laboratory testing. Rheumatology is as much at a loss as other specialties. Examples are given of important missed diagnoses revealed only at autopsy.When I was a medical school student in the 1950s and a resident and junior staff member in the 1960s and 1970s, regular visits to the morgue were an indispensable part of our workday routine. No first-class teaching hospital could operate without an adequate post-mortem service. The contact between clinicians and pathologists in the morgue was an essential part of ultimate quality control and teaching and could stimulate collaborative research. In the city of Malmö, where the autopsy rate exceeded 80% of the population in the 1970s, it is now less than 10%. When I asked a resident when he last had been to a morgue, he answered ‘I was there once when still a medical student’. Do the improved diagnostic instruments available today fully compensate for the lack of direct information given by careful autopsies, or are we missing valuable information by neglecting routine autopsy? All evidence indicates that we are indeed missing important information.In the 1970s, a 65-year-old man received treatment for systemic sclerosis on the basis of skin thickening, dysphagia, constipation with abdominal pain, and cardiomegaly. Two days before Christmas Eve he died of heart failure. The next day, the professor in the morgue greeted us with a big smile: ‘Today it is Christmas even here. I have today sectioned my first case of Chagas disease’. The patient, a teacher, had spent time in Colombia, a fact the clinicians had paid little attention to. The physician in charge of the patient did research in scleroderma and learned a lesson for life.Professor Kuntal Chakravarty, of Romford, UK, recently told me of a 36-year-old woman with a 5-year history of scleroderma who was admitted with acute abdominal pain, vomiting, and fever. X-rays and ultrasound did not reveal a cause. Owing to her scleroderma, the surgeons were reluctant to perform laparotomy, and she was treated with parenteral antibiotics and intravenous fluid. Her condition improved initially but later deteriorated and she died. The clinical diagnosis was peritonitis and ruptured intestine. The consulting rheumatologist (KC) and the family insisted on an autopsy, which unexpectedly showed uncomplicated volvulus.In 1975, we published an article on cause of death in 104 patients with rheumatoid arthritis (RA) based on routine post-mortem examination of patients during 5 years in a chronic care hospital in Malmö [1]. Cervical spine compression was identified as the cause of death in 11 patients. Only two of the cases had been diagnosed before death, although all patients had been hospitalized for months or years [1]. Although cervical spine instability is now rare, it still occurs and may be prevalent in communities with undeveloped health systems [2].Very high autopsy rates generated accurate prevalence studies of atherosclerosis [3], thromboembolism [4] and cancer [5,6]. In 1969, when the autopsy rate was 65%, Görel Östberg examined all 1,097 (!) temporal arteries from patients dying in 1 year in the city of Malmö, which had 250,000 inhabitants. The prevalence according to the literature was 2 out of 100,000, but Östberg identified not fewer than 16 out of 1,000 patients with a male/female ratio of 6/10. Only two of the patients had received a clinical diagnosis of temporal arteritis (retrospectively), and only a couple had suggestive symptoms [7]. Even more interesting is her systematic study of large vessel involvement in polymyalgia rheumatic and temporal arteritis, showing their overlap and coexisting polyarteritis nodosa and Takayasu’s disease [8]. This work has recently been fully acknowledged and extended by Gary S Hoffman and the US Vasculitis Clinical Research Consortium [9].In Finland, rheumatologists have published a number of articles illustrating substantial discrepancies between clinical and autopsy-based causes of death. In 36% of 371 autopsied patients, significant infections were identified, only half of which had been diagnosed in vivo[10]. Amyloidosis was the cause of death in 9.5% of patients between 1950 and 1991; of these cases, 35% had not been diagnosed in vivo[11]. Coronary heart disease showed an increasing prevalence in the same time period in the RA patients in contrast to autopsied non-RA patients and were likewise often not detected before death [12].In the US, the autopsy rate dropped from above 70% in the 1960s to 20% in 2005 at the Mayo Clinic and the Brigham and Women’s Hospital. The overall rate in the US is now 4.3% in non-forensic cases [13]. Although advances in diagnostic tools have improved accuracy of clinical diagnostics, 8% of major errors were found in a systematic review of 53 publications [14]. Post-mortem imaging has been investigated as an alternative to autopsy [15]. An ambitions blinded comparison between computed tomography (CT), magnetic resonance imaging (MRI), and autopsy of 182 cases reported to the coroner between 2006 and 2008 showed that CT was as accurate as the clinical diagnosis but that nevertheless causes of sudden death were often missed. MRI was less accurate than CT [16]. Even in the context of experimental medicine, the trend is the same. In the Autologous Stem Cell Transplantation International Scleroderma trial of stem cell transplantation versus cyclophosphamide in diffuse systemic sclerosis, autopsy was performed in 7 out of 44 cases (Jaap van Laar, of Newcastle, UK, and Kamran Naraghi, of Middleborough, UK).

Conclusions

The decline in autopsy rates has several causes: diagnostic overconfidence, physician reluctance to pursue consent for autopsy, lack of regulatory requirements, public resistance, and budgetary constraints. Part of the problem is that doctors are short on time and motivating the family to agree to autopsy requires tact, time, and dedication. Reluctance to embark on the additional paper work may also contribute. Autopsy remains the gold standard for determining the cause of death [13], and although revitalizing it seems unrealistic, I feel a need to alert younger generations what they are missing, in rheumatology as in other specialties.

Box 1. About Frank Wollheim

Frank Wollheim is Emeritus Professor at the Department of Rheumatology, University of Lund. He was its chairman from 1982 to 1998 and started its programs in scleroderma, systemic lupus erythematosus, early rheumatoid arthritis, and biomarkers of arthritis. He trained in internal medicine with Jan Waldenström in Malmö and rheumatology with Ralph C Williams in Minneapolis before starting the first rheumatology unit in Malmö in 1972. He has served as Secretary General and board member of Osteoarthritis Research Society International and is a master member of the American College of Radiology.

Abbreviations

CT: Computed tomography; MRI: Magnetic resonance imaging; RA: Rheumatoid arthritis

Competing interests

The author declares that he has no competing interests.